Loss of UBE3A expression in neurons leads to the neurodevelopmental disorder Angelman Syndrome (AS). Dysregulation of UBE3A has been implicated in a variety of other neurodevelopmental disorders including 15qdup syndrome, idic15 syndrome, epilepsy, seizures, Rett Syndrome, and idiopathic autism. UBE3A, located on chromosome 15q11-13, is imprinted specifically in neurons and mono-allelically expressed from the maternal allele. Investigating the mechanism of UBE3A imprinting and the role of the paternally expressed UBE3A-antisense transcript is critical for understanding the complex transcriptional control that occurs in this region. Imprinting of UBE3A is hypothesized to occur through the expression of an antisense transcript on the paternal allele that suppresses expression of UBE3A from the paternal allele. However, the process by which the expression of the antisense transcript silences the paternal allele in order to create mono-allelic, maternal expression is not known. To identify the mechanism that suppresses paternal allele expression, a novel cell culture system will be used to probe allele-specific differences on the maternal and paternal copies of UBE3A. Previously, attempts to differentiate maternal and paternal specific characteristics have relied on sequence differences between the two alleles. In this proposal, a hybrid mouse- human cell line containing a single human chromosome 15 in a mouse neuronal cell line will be utilized to distinguish the allele-specific chromatin marks of maternal and paternal human chromosome 15 without relying on genetic polymorphisms, allowing us to answer questions that previously evaded investigation. The goals of this project are to test a physical occlusion model for silencing the paternal allele of UBE3A, and to characterize the localization of a paternally expressed long non-coding antisense RNA from this chromosome region during neurodevelopment for its potential role in the UBE3A imprint mechanism and paternal-specific chromatin decondensation of this locus during neuronal maturation. This project has implications for understanding tissue-specific imprinting, particularly in neurodevelopment, which will contribute to our understanding of autism spectrum disorders, and may suggest approaches to counteract the maternal deficiency of UBE3A in Angelman Syndrome. PUBLIC HEALTH RELEVANCE: UBE3A is an epigenetically regulated gene on human chromosome 15q11-13 essential for neuronal maturation, and is implicated in Angelman Syndrome and other autism-spectrum disorders. There is currently a poor understanding of how UBE3A becomes exclusively maternally expressed specifically in mature neurons in the postnatal brain. This proposal seeks to identify the mechanism of UBE3A imprinting during neuronal maturation through novel experimental systems. The successful completion of this study should yield findings to help develop therapeutic interventions for Angelman Syndrome and possibly other autism-spectrum disorders.